Validation of a Low-Volume (100 μL) Plasma Protocol for HIV-1 RNA Quantification Using the Hologic Aptima HIV-1 Quant
Lorna S Madurai1, Kiveshan Reddy2, Someshni Nair2
1Universal Pathology Laboratory, Prospecton, South Africa; CAPRISA, Centre for the AIDS Programme of Research in South Africa, Durban, South Africa.
Insights
A new low-volume protocol for HIV-1 viral load testing in children uses only 100 μL of plasma. This method accurately quantifies HIV-1 RNA, improving testing accessibility for pediatric patients.
Area of Science:
- Virology
- Clinical Diagnostics
- Pediatric Infectious Diseases
Background:
- Pediatric HIV-1 viral load monitoring is challenged by limited blood volumes in infants and children.
- Standard assays require 0.5-0.7 mL plasma, posing significant collection difficulties.
Purpose of the Study:
- To evaluate a modified low-volume (100 μL) protocol for the Hologic Aptima HIV-1 Quant Dx assay.
- To determine if the low-volume protocol's performance is equivalent to the standard 700 μL protocol.
Main Methods:
- Analytical validation using de-identified EDTA plasma samples across a range of HIV-1 RNA concentrations.
- Comparison of the 100 μL and 700 μL protocols on the Panther system.
- Assessment of agreement using log10 differences, correlation, and Bland-Altman analysis; precision evaluated with replicate testing.
Main Results:
- The 100 μL protocol showed excellent concordance with the standard method (mean log10 difference ≈ 0.00, max difference 0.12 log10 copies/mL).
- No false-positive or false-negative results were detected in suppressed samples.
- Precision analyses demonstrated minimal variability within predefined acceptability criteria.
Conclusions:
- The low-volume Aptima protocol provides accurate and reliable HIV-1 RNA quantification.
- This validated method can expand access to viral load testing in pediatric populations and other settings with limited plasma volume.
Abstract:
Pediatric HIV-1 viral load monitoring is often limited by the small blood volumes that can be safely obtained from infants and young children. Standard assays typically require 0.5 to 0.7 mL of plasma, which can pose challenges in these settings. This study evaluated a modified low-volume protocol using 100 μL of plasma, diluted for use with the Hologic Aptima HIV-1 Quant Dx assay, to determine whether performance was equivalent to the standard 700 μL protocol. An analytical validation was conducted by using de-identified EDTA plasma samples spanning a wide range of HIV-1 RNA concentrations. Each specimen was tested by using both protocols on the Panther system. Agreement between methods was assessed by using log10 differences, correlation analysis, and Bland-Altman analysis; within-run and between-run precision was evaluated by using replicate testing of negative, low-positive, and high-positive samples. The 100 μL protocol displayed excellent concordance with the standard method, with a mean log10 difference of approximately 0.00 and a maximum difference of 0.12 log10 copies/mL. No false-positive or false-negative results were observed among 30 fully suppressed samples. Precision analyses showed minimal variability, with all results within predefined acceptability criteria. These findings indicate that the low-volume Aptima protocol provides accurate and reliable HIV-1 RNA quantification and can expand access to viral load testing in pediatric patients and other clinical settings in which plasma volume is limited.
